DESCRIPTION: TRIM5? restriction factors bind retroviral capsids after cell entry and restrict retroviral infection by blocking reverse transcription and/or integration of the viral genetic material. The C-terminal SPRY domain of TRIM5? is believed to form most of the capsid-TRIM5? interface as species-specific sequence variations within the SPRY domain account for differences in the viral specificity of the TRIM5?-mediated restriction. Most notably, recent evolution of the human TRIM5? SPRY resulted in the variant that has poor affinity for the HIV capsid, the vulnerability that contributed to the emergence of the AIDS pandemic when the simian immunodeficiency virus (SIV) passed from chimpanzees into a human host. Mutation or deletion of a single amino acid residue (R332) within the SPRY domain restores HIV restriction by the human TRIM5? variant. TRIM5? SPRY domains have proven refractory to structural and biophysical studies limiting our understanding of retroviral capsid recognition by TRIM5?. This research proposal seeks to fill that knowledge gap by solving the structures of several primate TRIM5? SPRY variants with distinct retroviral specificities and characterizing their interactions with the cognate retroviral capsids. The goal of the proposed research is to understand the mechanism of capsid recognition by TRIM5? restriction factors, elucidate why the human TRIM5? variant is not potent against HIV and to explore whether this defect could be alleviated by pharmacological or other means. The following three specific aims will be pursued: (1) The structures of the rhesus, human and possibly other TRIM5? SPRY domains will be solved using a novel experimental approach that combines NMR spectroscopy and X-ray crystallography. (2) Interaction of several SPRY-capsid pairs will be characterized using a variety of biophysical approaches. (3) High-resolution models of the SPRY-capsid complexes will be generated using computational methods and tested using mutagenesis and restriction assays.